In order to investigate how energy expenditure changes with over and underfeeding the following studies are underway. In one study, after careful calibration of weight maintenance EE, individuals undergo a series of measurements of 24 hour EE in a respiratory chamber in which they are fasting or overfed (by 200% of weight maintenance needs) a series of diets that vary in macronutrient content. This is to further investigate whether low or high protein diets may improve the detection of recruitment of adaptive thermogenesis. In addition, behavioral, metabolic and hormonal tests are performed to examine associated characteristics and to investigate the mechanism of the changes in EE. These individuals will also be followed up long term to look at what factors predict weigh change. We found that the change in energy expenditure with fasting and with overfeeding is reproducible. In addition in our initial results, the increase in energy expenditure with overfeeding was highest in those overfed a normal protein high fat or high carbohydrate diet. Energy expenditure did not increase significantly with low protein diets. We have now enrolled over 80 participants in this study. We have found a negative association between the percent change in EE with overfeeding with the percent change in EE with fasting indicating the presence of thrifty versus spendthrift phenotypes. Moreover, in short term (6 month) follow-up we found that changes in less energy expenditure decrease with fasting were associated with less weight gain. Based on the difference in energy expenditure measured during fasting and overfeeding, the thermic effect of food (TEF) can be calculated. We have found that TEF is inversely associated with adiposity. Thus TEF may predispose to weight gain or be an adaptation to increased bodyweight. As increased adiposity may insulate against trans-abdominal heat loss which may increase TEF, we are investigating the effect of central insulation on the EE and TEF changes associated with overfeeding. We are also continuing to measure EE with overfeeding and fasting, and investigating whether these changes predict longer term weight gain. Because of the recent discovery of the presence of brown fat in humans and its possible role in thermogenesis, we performed positron emission scans with labeled glucose. As brown fat is activated by cold temperatures, we have established that we can visualize brown fat after 2 hours of exposure to 16 degrees Celsius. We then currently investigated whether individuals with visualized brown fat after cold exposure, have visualized brown fat after overfeeding. Following demonstration of visible brown fat after cold exposure individuals were overfed by 200% of their energy needs using a high fat normal protein diet while in our metabolic chamber. The next morning they underwent a PET-CTscan; this was performed in some individuals prior to breakfast (approximately 12 hours after their last overfeeding meals) and in some individuals following a similar overfeeding breakfast (approximately 4 hours after their last meal). We found no evidence of activation of brown fat with overfeeding following a high fat overfeeding, indicating that brown fat does not mediate the increased energy expenditure associated with overfeeding. We are currently investigating whether brown fat might be activated following 24 hours of high carbohydrate overfeeding. In a related study, after measurement of EE with over and underfeeding, and also after undergoing a series of metabolic and behavioral testing (including biopsies of muscle and fat), individuals are admitted for 6 weeks of an inpatient dietary protocol involving underfeeding (for overweight and obese individuals) or overfeeding (for lean, obesity resistant individuals). During the inpatient study, all aspects of food intake, energy expenditure, and energy loss are carefully measured to determine if differences in weight gain or loss can be attributed to recruitment of adaptive thermogenesis or other factors. We have currently completed 12 individuals who have undergone the weight loss study. In these individuals, we found that the calculated energy deficit (based on measured loss of fat mass and fat free mass) did not significantly differ from the deficit as calculated based on energy intake, measured energy expenditure, and stool and urine losses. We did find that less decrease in 24hEE with fasting was associated with greater weight loss in this controlled inpatient study. Participants with the greatest accumulated measured energy deficit lost the most weight, and greater measured energy deficit was also associated with less 24h EE decrease with fasting. These results indicate the presence of spendthrift versus thrifty phenotypes even in obese individuals undergoing weight loss, and accounts in part for the variability in dietary weight loss. Recruitment for both studies is ongoing.